LAAN-A-SP-E009 No.B32 Microchip Electrophoresis Identification of Common Bean Cultivars by RAPD-STS with MCE-202 "MultiNA" In Japan, when a new variety of plant is breeded and its variety registration is accepted, the registered breeder of that variety is granted rights (breeder’s right) to that specific plant cultivar according to Japan's Plant Variety Protection and Seed Act. For example, kidney (common) beans are cultivated all over the world, and many varieties are in circulation. A domestic variety of the kidney bean (tebo) is used as the chief ingredient in sweet white bean paste. However, infringe problems against the breeder’s right occur. Without the permission of the breeder for the registered variety, the variety is exported and cultivated in overseas or the resultant unlicensed plant is re-imported to Japan. In conjunction with the visual inspection of the fruiting body, qualitative Polymerase Chain Reaction (PCR) genetic testing is effective for distinguishing among the common bean cultivars; the RAPD-STS (Random Amplified Polymorphic DNA Sequence Tagged Sites) method has been developed to accomplish this. Here we introduce cultivar identification of common beans by RAPD-STS method using the MultiNA. For the samples, we used two varieties of commercially available domestic common beans (Yukitebo and Gintebo), and two overseas varieties (pea bean and great northern bean). Cultivar identification was conducted according to the RAPD- STS method (Japanese Patent No. 2004-16008). With the RAPD method, random primers are used. However, a feature of the RAPD-STS method is that the primers are selected based on the sequence information of the obtained PCR products. The above- mentioned four varieties of common beans were pulverized and the DNA was extracted using the DNeasy Plant Mini Kit. The extracted DNA solutions were measured using the BioSpec-nano life science UV-VIS spectrophotometer, to determine the DNA concentrations. Then, DNA extract (10 ng) from each of the bean cultivars was used as a template, and three types of primers (SP01, SP02, SP03) were used for conducting the respective PCR amplifications. The PCR products were analyzed using the MultiNA, their appearance patterns were then determined (Fig. 1). ● DNeasy Plant Mini Kit (50) (QIAGEN) 69104 ● DNA-1000 Kit (Shimadzu) P/N 292-27911-91 ● SYBR ® Gold nucleic acid gel stain (invitrogen) S11494 ● 100 bp Ladder (Takara Bio) 3407A n Analytical Procedure n Reagents / Kits Analytical Instrument : MultiNA Analysis Mode : DNA-1000 on-chip mode n Analytical Conditions for PCR Products Extracted DNA PCR products Common bean cultivar identification PCR using primer set for identification Analysis of PCR products MultiNA Common bean Crushing, DNA extraction Collation of patterns Acquisition of appearance patterns of PCR products Nucleic acid quantitation BioSpec-nano Fig. 1 Procedure for Identification of Common Bean Cultivars